The COP9 signalosome (CSN) complex is an evolutionally conserved protein complex present in all eukaryotes and is involved in controlling diverse cellular and developmental processes. Disregulation of the CSN complex can have a dramatic effect on cellular functions critical to tumor development, including maintenance of DNA fidelity, cell cycle control, DNA repair, angiogenesis, and micro-environmental homeostasis. The CSN complex has recently been found to be involved in the regulation of NF-?B activated by excellular signaling. But a full understanding of the function and regulation of the CSN complex in signal-induced NF-?B activation pathways is lacking. NF-?B proteins are inducible transcription factors that are crucial regulators of many physiological and pathophysiological processes. Aberrant regulation of NF-?B and the signaling pathways (e.g. TNFa pathway) controlling its activity are involved in cancer development and progression, as well as resistance to chemotherapy and radiotherapy. Therefore, NF-?B and components involved in regulating its activity have become a focal point for drug discovery. Given its critica importance in cancer development, we hypothesize that detailed analysis of the CSN complex in the NF-?B pathway will not only provide new insights into its function and regulation in general, but also lead to new directions in future development of clinically useful inhibitors for cancer treatment targeting the NF-?B system through the CSN complex. To test this, we propose to achieve the following specific aims: 1) To define protein interaction dynamics of the human CSN complex associated with NF-?B activation triggered by TNFa signaling. 2) To characterize CSN5 phosphorylation and the dynamic interactions of the CSN5-IKK complex upon TNFa signaling. The proposed work represents the first detailed proteomic analysis of the CSN complex in the context of signaling pathways. The methodology developed in this work will be applicable for the study of other protein complexes in response to extracellular signaling. PUBLIC HEALTH RELEVANCE: Project Narrative Disregulation of the NF-?B pathway can lead to many human diseases including cancer. The CSN complex is a multi-subunit complex that is involved in regulating turmorigenesis and is associated with NF-?B activation. Detailed characterization of the CSN complex in the NF-?B pathway will allow the identification of potential molecular targets for improved cancer therapeutics.